Genomic Insights Into the Interspecific Diversity and Evolution of Mobiluncus, a Pathogen Associated With Bacterial Vaginosis

Abstract

Bacterial vaginosis (BV) is a common vaginal infection and has been associated with increased risk for a wide array of health issues. BV is linked with a variety of heterogeneous pathogenic anaerobic bacteria, among which Mobiluncus is strongly associated with BV diagnosis. However, their genetic features, pathogenicity, interspecific diversity, and evolutionary characters have not been illustrated at genomic level. The current study performed phylogenomic and comparative genomic analyses of Mobiluncus. Phylogenomic analyses revealed remarkable phylogenetic distinctions among different species. Compared with M. curtisii, M. mulieris had a larger genome and pangenome size with more insertion sequences but less CRISPR-Cas systems. In addition, these two species were diverse in profile of virulence factors, but harbored similar antibiotic resistance genes. Statistically different functional genome profiles between strains from the two species were determined, as well as correlations of some functional genes/pathways with putative pathogenicity. We also showed that high levels of horizontal gene transfer might be an important strategy for species diversification and pathogenicity. Collectively, this study provides the first genome sequence level description of Mobiluncus, and may shed light on its virulence/pathogenicity, functional diversification, and evolutionary dynamics. Our study could facilitate the further investigations of this important pathogen, and might improve the future treatment of BV.

Keywords: Mobiluncus; bacterial vaginosis; comparative genomics; horizontal gene transfer; interspecific divergence; pathogenicity.

FIGURE 1

Genomic features of the genus Mobiluncus. (A) Hierarchical clustering based on ANI values of 38 Mobiluncus strains. (B) Whole genome alignment of M. curtisii and M. mulieris. Color denotes percent similarity of links (see legend). (C–J) Comparison of genomic characteristics between M. curtisii and M. mulieris. The boxplot shows the median, and the first and third quartiles as the lower and upper hinges. Outliers are indicated as dots. Asterisks (***) indicate significant differences (significance level of 0.001, Wilcoxon test). NS, not significant; ISs, insertion sequences; CRISPR, clustered regularly interspaced short palindromic repeat; ARGs, antibiotic resistance genes; VF, virulence factor.

FIGURE 2

Phylogeny of the genus Mobiluncus. (A) Maximum-likelihood phylogenomic tree. The tree was constructed based on 329 core genes that could give a well-resolved tree topology without recombination. The total numbers of ancestral orthologous genes present at each of the nodes are shown next to the deep nodes, and the numbers of gene gain (+) and loss (–) events are indicated below. Bootstrap values less than 70% are shown at the nodes. The scale bars indicate 10 and 0.25% sequence divergences among and within species, respectively. (B) Hierarchical cluster analysis based on the presence or absence of dispensable genes. Height indicates the dissimilarity between genomes. BP (bootstrap probability) values less than 70% from 1,000 replicates are shown, and all AU (approximately unbiased) p-values are > 70%.

FIGURE 3

Hierarchically clustered heatmaps of the distributions of the putative antibiotic resistance genes (A) and virulence factors (B) in Mobiluncus genomes.

FIGURE 4

Pangenome summary statistics of M. curtisii and M. mulieris. (A) Histogram distributions of soft-core, shell, and cloud genes. Pie chart displays percentages of each part of the total genes. (B) The sizes of pan- and core-genomes in relation to numbers of genomes added into the gene pool.

FIGURE 5

Functional divergences between M. curtisii and M. mulieris. (A) Boxplot of abundance of differential COG categories. All the distributions were significantly different (Wilcoxon test, p < 10-5). (B) Heatmap of COGs that significantly contributed most to the dissimilarity between M. curtisii and M. mulieris. Different COG categories are shown in different colors.

FIGURE 6

Genetic organizations of the regions containing species-specific genes. Only genes that are universal (> 90%) in one species but absent in the other are determined as species-specific genes. Conserved genomic regions flanking these genes are colored in gray. Non-specific genes are marked with dashed lines. MoCo, molybdenum cofactor.

FIGURE 7

Analysis of predicted transferred genes. (A) Distribution of the predicted transferred genes in soft-core, shell, and cloud genes. Numbers of transferred genes and gene families are indicated. (B) Pie chart displays percentages of different donors of HGT events at order level. (C) Non-metric multidimensional scaling (NMDS) plot of COG entries showing distinct clustering of each species.